CORE AREA 1 - CHRONIC DIABETES -

CASE E

Mrs HT, 60 years, has essential hypertension & NIDDM.

Regular Medications Taken

• Felodipine 10mgs BD

• Atenolol 50mgs mane

• Chlorothiazide 500mgs mane

• Metformin 500mgs TDS

A recent 24 hour urine shows:

Urine albumin 50 mgs

(Normal range is less than 15 mgs in 24 hours)

Discussion:

What is the relevance of the clinical chemistry result for the diabetic patient especially with respect to the possibility of long term complications.

RELEVANCE

• Poor control of diabetes is associated with enlargement of the kidneys & initially a high GFR, which are early functional abnormalities in diabetic nephropathy.

• Microalbinuria is a marker of renal damage in the diabetic state.

• Patients with microalbinuria over time develop progressive renal injury.

Microalbinuria

• A marker of the development of proteinuria• Early mortality in NIDDM patients• Associated with unfavorable lipid profiles• Increasing blood pressure elevation• Retinopathy• Untreated it is progressive• Progressive decline in GFR• Higher level of structural damage present

Microalbinuria

• Poor metabolic control is a risk factor for the development and progression of microalbinuria as well as a decline in GFR

• Diabetic nephropathy progresses over 10 to 25 years, it is the most common cause of end stage renal disease in the USA

• Diabetic nephropathy ultimately presents as the nephrotic syndrome

The Nephrotic Syndrome

• Results from alterations in the permselective characteristics of the Glomerular Basement Membrane (caused by hyperglyceamia), that allows increased passage of proteins of intermediate size into the urine

The Nephrotic Syndrome

• Consists of heavy proteinuria, hypoalbuminemia, hyperlipidemia, higher concentration of high MW proteins & oedema formation, associated with hypertension.

Methods used to measure urine albumin level.

Methods to measure urine albumin level

Quantitative assay– Radioimmunoassay– Immuno-nephelometry– Electro-immuno-diffusion assay– Immunoturbidimetric assay

Semi-quantitative assay- test strip

Methods to measure urine albumin level

*Radioimmunoassay-Sensitive,precise and rapid method-Reliable results

*Immuno-nephelometry and Electro-immuno-diffusion assay

-sensitive and accurate-correlated with RIA

Methods to measure urine albumin level

*Immunoturbidimetric assay

-for initial screening of albuminuria

*Semi-quantitative method

-test strip

-high sensitivity but poor specificity

-use of this method questionable

Drugs that can affect urine albumin level

• Acetazolamide• Aminoglycosides• Amphotericin• Cephalosporins• Colistin• Griseofulvin• Lithium• Nephrotoxic drugs eg arsenicals, gold salt• Penicillamine

Drugs that can affect urine albumin level

• Penicillin G

• Polymyxin B

• Salicylates

• Sulfonamides

• Tolbutamide

Disease factors that could affect urine albumin level

• Bacterial pyelonephritis

• Bladder tumor

• Congestive heart failure

• Diabetic nephropathy

• Glomerulonephritis

• Heavy metal poisoning

• Lupus erythematosus

Disease factors that could affect urine albumin level

• Malignant hypertension

• Multiple myeloma

• Nephrotic syndrome

• Nephrotoxic drug therapy

• Polycystic kidney disease

• Preeclampsia

• Urinary tract infection

Other interfering factors for urine albumin

• Severe emotional stress

• Strenuous exercise

• Urine contamination with secretion

Advanced Glycation Endproducts

(AGE)

What Are AGEs?• A class of complex products, brown in colour

and with specific fluorescence• Very unstable, reactive compounds and hence

difficult to analyse completely• The result of a succession of chemical reactions

linked in a complex network (known as the Maillard reaction)

• Binding of AGEs to RAGE (AGE receptors) activates endothelial cells and monocytes to produce cytokines, adhesion molecules and tissue factors.

AGE Production

• Non-enzymatic glycosylation between a free amino group on a protein and a carboxyl group from a sugar e.g. glucose to form Schiff’s base

• Schiff base formation is relatively fast and highly reversible and is an equilibrium reaction. (The amount of Schiff base formed is dictated by glucose concentration.)

• Over 24-48 hours Schiff’s base undergoes molecular rearrangement to create an N-substituted glycosylamine

• This leads to formation of more stable Amadori products (1-amino-1-deoxy-2-ketose)

How are they formed? Continued….

• The formation of the Amadori product from the Schiff base is slower, but much faster than the reverse reaction and therefore tends to accumulate on proteins. (Equilibrium levels of Amadori products are reached in weeks)

• As with the formation of the Schiff base, the amount of Amadori product formed is related to the glucose concentration.

• On long lived proteins Amadori products rearrange further to form AGEs

The formation of advanced glycosylation end-products (AGE).

• Best chemically characterised AGE compounds found in humans are pentosidine and carboxyl methyl lysine.

• AGES may also originate from arabinose and xylose from food, e.g., intestinal bacteria degrade xylanes found in fruit to produce AGEs

• AGEs are toxic, inducing bacterial mutagenesis

• Formed in excess in diabetes mellitus, renal failure and the elderly.

AGE and RAGE (Receptor for AGE)

• RAGE receptors are expressed on a wide range of cells including smooth muscle cells, monocytes, macrophages, endothelial cells and astrocytes.

• The RAGE receptor is thought to mediate a complex pattern of intracellular signalling.

• RAGE expression is increased in the blood vessels and kidneys of diabetic patients.

What effect do AGEs have on the body?

Relevance to Diabetes.

• AGEs are believed to contribute to the chronic complications (Secondary complications) of diabetes mellitus.

• Which include: Visual Impairment, Blindness (Retinopathy), Renal Failure (Nephropathy), Stroke, MI, Neuropathy and other Cardiovascular Complications.

Relevance to Diabetes.

(Nephropathy)

• NEJM conducted a study to measure serum and tissue AGEs in diabetic (Types I and Type II) and nondiabetic patients with different levels of renal function.

• The mean AGE content of samples of arterial-wall collagen from 9 diabetic patients was significantly higher than that of samples from 18 nondiabetic patients.

Relevance to Diabetes.

(Nephropathy) Continued …..

• In diabetic patients: The levels of AGE peptides correlated directly with serum creatinine and inversely with creatinine clearance, suggesting that levels of AGE peptides increased with the severity of diabetic nephropathy.

• Conclusion: AGEs accumulate at a faster-than-normal rate in arteries and the circulation of patients with diabetes; the increase in circulating AGE peptides parallels the severity of renal functional impairment in diabetic nephropathy

Relevance to Diabetes (Vascular)

• Initial AGE- RAGE interaction- cellular activation and inflammation.

• Followed by lipoprotein accumulation and atherosclerosis.

• AGE also depletes nitric oxide.

• Resulting in vascular thickening with loss of elasticity and hypertension.

Relevance to Diabetes (Vascular) Continued….

• Schematic description of the formation of collagen cross-links. AGEs can react with free amino groups on an adjacent protein to form cross-links. Cross-linking can lead to decreased compliance of large vessels and of the myocardium

Relevance to Diabetes (Retinopathy)

• Characterised by abnormal vessel development leading to haemorrhages.

• Proposed that AGE may increase retinal endothelial call permeability resulting in vascular leakage.

Conclusion.

• Measuring the concentration of serum AGE’s is becoming more important as studies are concluding that they are useful as a marker for or indication of possible secondary diabetic complications.

Haemoglobin A1C. (Also known as Glycosylated Hb)

• An amadori glycoylation product.• Is the stable product of nonenzymatic glycosylation of the

B- chain of Hb by plasma glucose. • Indicator of plasma glucose control over the previous 1-3

months.• Normal levels is around 6%. • In poorly controlled diabetics level ranges from 9- 12%.• HbA1c is not a specific test for diabetes but often indicates

existing diabetes.• Often viewed as a measure of long term control.

Fructosamine

• Because the turnover rate of proteins is much faster than that of hemoglobins, the measurement of glycated serum proteins (i.e., fructosamines) provides a shorter term, more rapidly responding assessment of diabetic control. The fructosamine measurement reflects an average blood glucose level of the past 2-3 weeks, while GHb values indicate control over a 2-3 month period. Fructosamine levels are directly proportional to serum protein levels (albumin, immunoglobulins). Since the fructosamine level is not given as a percentage of actual serum total protein, a low level may indicate good control or it may indicate low total protein. Variations in levels of serum proteins (albumin, IgA, and other immunoglobulins) may affect fructosamine results.

• Non-diabetic patient: 0-285 µmol/L

References

-         Australian Medicines Handbook 2002 – Simone Rossi, Agnes Vitry, Eve Hurley, Fay Abbott, Sharon Goldsworthy, Christopher Alderman, Manya Angley, Neil Hotham. Finsbury Press 2002.-         Diseases of the Kidney 6th Edition Volume 1 – Robert W Schrier, Carl W Gottschalk. Little, Brown and Company 1997.-         Nephrology Volume 1 – Jean Hamburger, G Richet, J Crosnier, J L Funck-Brentano, B Antoine, H Ducroit, J P Mery and H de Montera. W. B. Saunders Company 1968.-         Papper’s Clinical Nephrology 3rd Edition – Francisco Llach. Little, Brown and Company 1993.-         Renal Pathology 2nd Edition – Frederick Dische. Oxford Medical Publications 1995.-         The Merck Manual 17th Edition – Mark H Beers, Robert Berkow. Merck Research Laboratories 1999.

Therapeutic Guidelines Antibiotics Version 11 – J Spicer. Therapeutic Guidelines Limited 2002.